βENaC is a molecular component of a VSMC mechanotransducer that contributes to renal blood flow regulation, protection from renal injury, and hypertension

Abstract

Pressure-induced constriction (also known as the "myogenic response") is an important mechano-dependent response in certain blood vessels. The response is mediated by vascular smooth muscle cells (VSMCs) and characterized by a pressure-induced vasoconstriction in small arteries and arterioles in the cerebral, mesenteric, cardiac, and renal beds. The myogenic response has two important roles; it is a mechanism of blood flow autoregulation and provides protection against systemic blood pressure-induced damage to delicate microvessels. However, the molecular mechanism(s) underlying initiation of myogenic response is unclear. Degenerin proteins have a strong evolutionary link to mechanotransduction in the nematode. Our laboratory has addressed the hypothesis that these proteins may also act as mechanosensors in certain mammalian tissues such as VSMCs and arterial baroreceptor neurons. This article discusses the importance of a specific degenerin protein, β Epithelial Na(+) Channel (βENaC) in pressure-induced vasoconstriction in renal vessels and arterial baroreflex function as determined in a mouse model of reduced βENaC (βENaC m/m). We propose that loss of baroreflex sensitivity (due to loss of baroreceptor βENaC) increases blood pressure variability, increasing the likelihood and magnitude of upward swings in systemic pressure. Furthermore, loss of the myogenic constrictor response (due to loss of VSMC βENaC) will permit those pressure swings to be transmitted to the microvasculature in βENaC m/m mice, thus increasing the susceptibility to renal injury and hypertension.

Keywords: baroreflex; degenerin; epithelial Na+ channel; hypertension; mechanotransduction; myogenic constriction; renal blood flow; renal injury.

Figure 1

Initiation of the myogenic response. (A) The sequence of events leading myogenic constriction. (B) Hypothetical role of degenerin proteins as a mechanotransducer in VSMCs.

Figure 2

Localization of βENaC and γENaC in enzymatically dissociated renal VSMCs. VSMCs were identified by labeling with a smooth muscle actin (left column). We detected βENaC and γENaC, but not αENaC, at or near the VSMC surface. The strongest labeling is consistently observed for βENaC. Scale bar represents 5 μm. (Figure includes images reproduced from American Journal of Physiology, Renal Physiology 289; F891–F901, 2005, Figure 2).

Figure 3

In vitro assessment of myogenic constriction. (A) Image of an isolated renal interlobar artery segment tied to inflow and outflow perfusion pipettes. (B) To assess myogenic responsiveness vessels are exposed to a step-wise increase in perfusion pressure (25 mm Hg step, 5 min each) from 25 to 150 mm Hg under normal external Ca²⁺ and Ca²⁺ free solutions to determine active response and passive responses, respectively (C). (C) The steady state diameter is plotted vs. pressure to obtain a pressure-diameter relationship under active vs. passive conditions. (D) Myogenic tone is calculated as [(passive diameter – active diameter)/passive diameter] at each pressure step. A vessel with active myogenic tone will have an increase in tone with an increase in pressure. A vessel with reduced or absent myogenic tone will have a flattened relationship between pressure and tone. (E) Effect of ENaC inhibition with benzamil on myogenic tone in renal interlobar arteries. (F) Transient gene-silencing of βENaC or γENaC using dominant-negative constructs of βENaC or γENaC inhibits myogenic tone in renal interlobar arteries. (G) Transient gene-silencing of βENaC or γENaC using siRNA inhibits myogenic tone in renal interlobar arteries. (Figure reproduced from American Journal of Physiology, Renal Physiology 289; F891-F901, 2005, Figure 4 and American Journal of Physiology, Renal Physiology 291; F1184–F1191, 2006, Figures 5, 6).

Figure 4

Localization of βENaC and contribution to myogenic constriction in renal afferent arteriole. (A) Localization of SM α-actin (left) and βENaC (right) in a cross-sectional image of a renal arteriole from a βENaC +/+ (top) and m/m (bottom) mouse. In the merged image, an “L” identifies the arteriolar lumen and an asterisk (^*) identifies the VSMC cell bodies. (B) Image of an isolated afferent arteriole with attached glomerulus preparation. (C) Steady state vasoconstrictor responses to an increase in pressure in afferent arterioles in βENaC +/+ and m/m animals (n = 6). Vasoconstrictor responses to adrenergic agonist norepinephrine (NE) were similar (not shown). (Figure reproduced from American Journal of Physiology, Renal Physiology 302; F1486–F1493, 2012, Figures 1, 2).

Figure 5

βENaC mediates myogenic regulation of whole kidney blood flow and renal vascular resistance. (A) Schematic of animal preparation. Mice have an arterial carotid catheter for blood pressure measurement and a renal flow probe for whole kidney blood flow measurement. An occluder tie placed around the lower abdominal aorta is used to generate the step increase in pressure. (B) Time course of the regulatory response of mean arterial pressure (MAP; B), renal blood flow (RBF; C), and renal vascular resistance (RVR; D), in wild type (+/+, filled symbols, n = 6) and mutant (m/m, open symbols, n = 7) 10 s before and 10 s after the step increase in pressure. Data are presented as normalized changes in B–D to minimize variance. Following a similar increase in MAP, the transient increase in RBF and decrease in RVR are similar between +/+ and m/m mice. Immediately following the transient drop, RVR begins to increase in the +/+, but remains low in the m/m. (E) The rate of increase in RVR during the first 5 s following the drop in RVR (Slope RVR_{0–5 S}) is significantly greater in +/+ vs. m/m mice (p = 0.014). By 20–30 s following the step increase in MAP, RBF in the +/+ is corrected while RBF remains elevated in m/m animals. By 20–30 s following the step increase in MAP, the change in RVR from baseline is greater in the +/+ vs. m/m. Data are mean ± SEM. ^*Significantly different from βENaC +/+ group at the p-value indicated. (Figure reproduced from American Journal of Physiology, Renal Physiology 302; F1486–F1493, 2012, Figure 3).

Figure 6

Blood pressure and arterial baroreflex gain in normal salt (0.4% Na⁺) fed animals. Mean arterial pressure (MAP) for 12-h light (L)–dark (D) cycles for each of 5 days and the average of 5 days are shown in panels A and B, respectively in wildtype (+/+, n = 8) and homozygous βENaC mutant mice (m/m, n = 7). (C) Arterial baroreflex gain is significantly lower in βENaC m/m mice (n = 4) vs. +/+ controls. (D) Mean arterial blood pressure (MAP) variability during the last 24 h of blood pressure recording was significantly elevated in βENaC m/m mice (same animals as shown in A and B). (Figure reproduced from American Journal of Physiology, Renal Physiology 301; F443–F449, 2011, Figures 2, 3). Data are mean ± SEM. BPM is heart rate in beats per minute. ^*Significantly different from wildtype (+/+) control animals, p < 0.05.

Figure 7

Schematic of working hypothesis: the role of βENaC as a mechanosensor and its role in cardiovascular pathophysiology. We hypothesize that βENaC is a critical component of mechanosensors in renal VSMCs and arterial baroreceptor neurons. Loss in βENaC function in renal VSMCs leads to a loss in renal myogenic constriction and a delayed or impaired correction of renal blood flow following upward swings in blood pressure. Loss in βENaC function in arterial baroreceptor neurons leads to a loss in baroreflex control of blood pressure and increased BP lability. Increased BP lability and decreased myogenic constriction increase the opportunities and magnitude of systemic pressure transmission to the renal microvessels, which in turn, increases susceptibility to end-organ injury and hypertension.